Polyketides, non-ribosomal peptides and other related natural products cover a region of chemical space that interacts very effectively with targets in biological systems, leading to very high hit rates on HTS and phenotypic screens (Koehn et al., 2005, Koehn, 2008, Carter, 2011). They have also been very successful commercially, with around 50 approved products with peak sales of the six most successful totalling $15 billion. The molecular diversity of polyketides and non-ribosomal peptides is very high, with macrocyclic, linear, mixed polyketide/peptide and glycosylated examples. For example, just over 7000 known polyketide structures have led to >20 commercial drugs. This 0.3% ‘hit rate’ compares very favourably with the <0.001% hit rate for synthetic compound libraries (Li and Vederas 2009). However, it is getting increasingly difficult to discover new natural product chemotypes from natural sources and new methods for increasing this ‘naturally available’ diversity are required.
Although polyketides are structurally diverse, they are produced by a common biosynthetic pathway. These pathways involve large enzymes, containing multiple modules each involved in one (or more) rounds of chain extension with Ketosynthase (KS), Acyl Transferase (AT) and Acyl Carrier Protein (ACP) domains with optional Dehydratase (DH), Enoyl Reductase (ER) and Ketoreductase (KR) (and sometimes other, such as methylase) domains. The polyketides are assembled in the producer organism by stepwise condensation of carboxylic acids (see Staunton et al., 2001 for review) followed by potential cyclisation and further processing of the beta-ketone function in a manner analogous to fatty acid biosynthesis, and generally exhibit a direct one to one correspondence between the genes encoding the polyketide synthase (PKS), the active sites of the biosynthetic proteins, the chemical reactions performed and the structure of the product molecule. Bioengineering techniques have been used to alter the genetic architecture coding for production of the PKS that generates the polyketide. However, the majority of previously described bioengineering techniques are only effective at making simple structural changes to the molecular structure of the parent polyketide (see Reeves et al., 2009 for review). A single genetic alteration leads to a specific chemical change in the polyketide produced. This is useful for lead optimisation and improvement of properties to make the polyketide of interest more drug-like, but has not been very successful at increasing the chemical space that naturally available polyketides cover, especially for generating new chemotypes.
Non-ribosomal peptides are produced by non-ribosomal peptide synthetases (NRPS), multimodular assembly lines which are analogous to polyketide synthases (see FIG. 1). In place of the KR, AT and ACP domains found in PKSs, there are Condensation, Adenylation and Thiolation (or Peptidyl Carrier Protein) domains (see Strieker et al., 2010 for review). Similar issues have been faced to those in PKS engineering (Giessen et al., 2012).
Thus, there remains a need to discover methods for more efficiently accessing novel analogues of natural products, in particular with significant alterations in gross structure.